AUTOIMMUNE DIABETES: EARLY EVENTS IN ISLETS OF LANGERHANS

自身免疫性糖尿病：朗格汉斯岛的早期事件

• 批准号: 9197630
• 负责人: EMIL Raphael UNANUE
• 金额: $ 45.57万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-08-01 至 2020-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9197630
• 关键词: Acute; Address; Alleles; Antibodies; Area; Autoimmune Diabetes; Beta Cell; Binding; Blood; CD4 Positive T Lymphocytes; CRISPR/Cas technology; Cells; Charge; Chemotactic Factors; Chimera organism; Clinical; Complex; Defect; Dendritic Cells; Development; Diabetes Mellitus; Disease; Epitopes; Event; Experimental Models; Fatty acid glycerol esters; Flushing; Gatekeeping; Gene Expression Profile; Genetic; Genome; Grant; Human; ITGAX gene; Image; In Vitro; Inbred NOD Mice; Insulin; Insulin-Dependent Diabetes Mellitus; Investigation; Islets of Langerhans; Lymphocyte; Lymphoid; Migration Assay; Minor; Modeling; Monoclonal Antibodies; Movement; Mus; NOR Mouse; Pancreas; Peptide/MHC Complex; Peptides; Phase; Process; Publishing; Reagent; Recruitment Activity; Reporter; Reverse Transcriptase Polymerase Chain Reaction; Role; Site; T-Lymphocyte; Testing; Thymus Gland; Time; Tissues; Translating; XCL1 gene; XCR1 gene; autoreactivity; base; chemokine; chemokine receptor; congenic; imaging approach; in vivo; islet; lymph nodes; migration; public health relevance; receptor; transcription factor

 DESCRIPTION (provided by applicant): This proposal is based on a recently published study showing that a minor subset of dendritic cells, the CD11c+CD103+ DC, those under the control of the Batf3 transcription factor, are absolutely required for diabetes development in NOD mice. These DCs are found both in islets and pancreatic lymph nodes and act as gate-keepers for diabetes initiation. Our aims center on discovering the processes whereby CD103+DC localize in islets. We combine examination of chemokines, antibodies, and live imaging with new preliminary findings that point to NOR mice having a defect in islets CD103+ DC, but not in pLNs. These areas of emphasis may point to strategies that may be translated to human T1D. Aim 1 investigates the mechanism of CD103+ DC entry into islets. The role of chemokines in CD103+ DC recruitment will be examined both in vitro and in vivo. Chemokine blockade will be tested in vivo. In vitro migration assays will define chemokines with the potential to recruit NOD-derived CD103+ DC to islets. RT- PCR will be used to determine which chemokines exist in islets of NOD mice at different times. Aim 2 will generate and examine several reagents intended to deplete or trace CD103+ DC in the NOD mouse. We have just generated an Xcr1-/- mice using Crispr/Cas technology which should bring definitive information on the role of this chemokine receptor in the islet localization of the CD103+ DCs. We will follow the same approaches used to examine the Batf3-/- mice just published. In a second subaim, a NOD.Xcr1-mOrange reporter mouse will be generated and used to trace the movement of CD103+ DC in the pancreas using live imaging approaches. We want to examine the traffic of CD103+DC in the pancreas and determine whether it may be the cell that transmigrates to the pLN. Finally an anti-Xcl1 monoclonal antibodies will be tested for its effects on CD103+ DC migration and protection from diabetes. Aim 3 takes advantage of our recent finding that NOR mice, which share 88% of their genome with NOD, lack the intra islet CD103+ DC but have them in LNs. By comparing the Batf3-/- mice with the NORs we expect to understand the mechanisms controlling early entrance of cells into islets. We examine this strain, and substrains containing NOR-derived alleles, to address the genetic behind the CD103+ DC entry into islets.

 描述（由申请人提供）：该提案基于最近发表的一项研究，该研究表明，树突状细胞的一小部分，即 CD11c+CD103+ DC，受 Batf3 转录因子控制，是 NOD 小鼠患糖尿病所绝对必需的这些 DC 存在于胰岛和胰腺淋巴结中，充当糖尿病发生的看门人，我们的目标是发现 CD103+DC 在胰岛中定位的过程。将趋化因子、抗体和实时成像检查与新的初步发现相结合，这些发现表明 NOR 小鼠在胰岛 CD103+ DC 中存在缺陷，但在 pLN 中没有缺陷。这些重点领域可能指向可转化为人类 T1D 的策略。研究 CD103+ DC 进入胰岛的机制 将在体外和体内测试趋化因子在 CD103+ DC 招募中的作用。体外迁移测定将确定具有将 NOD 衍生的 CD103+ DC 募集到胰岛的趋化因子，RT-PCR 将用于确定不同时间 NOD 小鼠胰岛中存在哪些趋化因子。或在 NOD 小鼠中追踪 CD103+ DC 我们刚刚使用 Crispr/Cas 技术生成了 Xcr1-/- 小鼠，这应该会带来有关该趋化因子受体在趋化因子受体中的作用的明确信息。 CD103+ DC 的胰岛定位。我们将遵循用于检查刚刚发布的 Batf3-/- 小鼠的相同方法，在第二个子目标中，将生成 NOD.Xcr1-mOrange 报告小鼠并用于追踪 CD103+ DC 的运动。我们希望使用实时成像方法检查胰腺中 CD103+DC 的流量，并确定它是否可能是迁移到 pLN 的细胞。将测试抗 Xcl1 单克隆抗体对 CD103+ DC 迁移和预防糖尿病的影响，Aim 3 利用了我们最近的发现，即 NOR 小鼠的基因组与 NOD 相同，但缺乏胰岛内 CD103+ DC。通过将 Batf3-/- 小鼠与 NOR 进行比较，我们希望了解控制细胞早期进入胰岛的机制。 NOR 衍生等位基因，用于解决 CD103+ DC 进入胰岛背后的遗传问题。